Exploring Phytochemical Mechanisms in the Prevention of Cholesterol Dysregulation: A Review

Although cholesterol plays a key role in many physiological processes, its dysregulation can lead to several metabolic diseases. Statins are a group of drugs widely used to lower cholesterol levels and cardiovascular risk but may lead to several side effects in some patients. Therefore, the development of a plant-based therapeutic adjuvant with cholesterol-lowering activity is desirable. The maintenance of cholesterol homeostasis encompasses multiple steps, including biosynthesis and metabolism, uptake and transport, and bile acid metabolism; issues arising in any of these processes could contribute to the etiology of cholesterol-related diseases. An increasing body of evidence strongly indicates the benefits of phytochemicals for cholesterol regulation; traditional Chinese medicines prove beneficial in some disease models, although more scientific investigations are needed to confirm their effectiveness. One of the main functions of cholesterol is bile acid biosynthesis, where most bile acids are recycled back to the liver. The composition of bile acid is partly modulated by gut microbes and could be harmful to the liver. In this regard, the reshaping effect of phytochemicals on gut microbiota has been widely reported in the literature for its significance. Therefore, we reviewed studies conducted over the past 5 years elucidating the regulatory effects of phytochemicals or herbal medicines on cholesterol metabolism. In addition, their effects on the recomposition of gut microbiota and bile acid metabolism due to modulation are discussed. This review aims to provide novel insights into the treatment of cholesterol dysregulation and the anticipated development of natural-based compounds in the near and far future.


INTRODUCTION
Many organizations have provided definitions of metabolic syndrome (MetS), the meaning of which has been progressively developed to be as comprehensive as possi-ble�in chronological order, these are the World Health Organization (WHO), in 1998; the European Group for the Study of Insulin Resistance (EGIR), in 1999; the National Cholesterol Education Program Adult Treatment Panel III (NCEP ATP III), in 2001; and the International Diabetes Federation (IDF), in 2005. 1 Among them, the NCEP ATP III definition provides one of the most widely used sets of criteria for MetS since it includes the crucial features of hyperglycemia/insulin resistance, visceral obesity, atherogenic dyslipidemia, and hypertension. 2According to this panel, in MetS, three or more of the following five criteria must be met: waist circumference over 40 in.(men) or 35 in.(women), blood pressure over 130/85 mmHg, fasting triglyceride (TG) level over 150 mg/dL, fasting high-density lipoprotein (HDL) cholesterol level less than 40 mg/dL (men) or 50 mg/dL (women), and fasting blood sugar level over 100 mg/dL. 2 Most of the above-mentioned criteria are correlated with the incidence of hyperlipidemia, a health condition characterized by the presence of excess plasma lipids (cholesterol, triglycerides, phospholipids, etc.) and lipoproteins (high-, low-, and very-low-density lipoproteins). 3 Hyperlipidemia is subclassified into hypercholesterolemia and hypertriglyceridemia, where an increase in plasma triglyceride levels is notably related to hypercholesterolemia.Although cholesterol is an essential precursor for biological molecules, including steroid hormones, bile acids, oxysterols, and vitamin D, the accumulation of free cholesterol has adverse effects on metabolism and health. 4A low-saturated-fat diet is always advised as a strategic approach to lower plasma cholesterol. 5owever, due to the inadequate cholesterol-lowering effects achieved from following dietary recommendations alone, statins (including pravastatin, simvastatin, atorvastatin, and rosuvastatin) have been employed and are efficacious in reducing plasma cholesterol, especially low-density lipoprotein (LDL) cholesterol. 5Nevertheless, statin drugs may lead to adverse effects in patients using multiple medications, including muscle damage, renal failure, and myopathy; in addition, scholars have suggested some risk factors associated with statin toxicity, such as alcohol abuse, frailty, multisystem diseases, gender, and age. 6Many preclinical and clinical studies have reported that nutraceuticals and phytochemicals from plant-based products exhibit cholesterol-lowering properties and can be potentially used as alternative medication for lowering the risks of atherosclerosis-related diseases. 7,8Therefore, researchers have encouraged the development of plantbased health products, including health food, functional food, and dietary supplements, containing nutraceuticals with hypocholesterolemic activity (as proven by their pharmacological effects) as a sustainable intervention to retard cholesterol increase. 9This review focuses on recent strategies and approaches to manage hypercholesterolemia using phytochemicals within 2019−2023.

REGULATION OF CHOLESTEROL HOMEOSTASIS
In a recent review, Duan et al. (2022) summarized the key steps and molecular mechanisms involved in the regulation of cholesterol homeostasis�including cholesterol biosynthesis, uptake, transport, utilization, and excretion�and the effect of epigenetic modulation on cholesterol metabolism. 10The subsequent sections focus on phytochemicals or plant-based products exhibiting regulatory effects on the molecular mechanisms of cholesterol homeostasis; most of the studies discussed in this review have been published between 2020 and 2023.

Effect of Phytochemicals on
Cholesterol Biosynthesis.The inhibition of cholesterol synthesis by phytoestrogens and phytochemicals has been suggested as a strategy to reduce blood cholesterol. 11This section discusses the regulatory effect of these natural compounds on the enzymes involved in controlling cholesterol biosynthesis, including sterol regulatory element-binding proteins (SREBPs), 3hydroxy-3-methylglutaryl-CoA synthase 1 (HMGCS1), 3hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR), and farnesyl-diphosphate farnesyltransferase 1 (FDFT1; Figure 1 and Table 1).
2.1.1.Sterol Regulatory Element-Binding Proteins (SREBPs).SREBPs regulate several downstream genes, including those for fatty acid synthase (Fasn), glycerol-3phosphate acyltransferase (Gpam), stearoyl-CoA desaturase (Scd), and proprotein convertase subtilisin/kexin type 9 (Pcsk9). 12The inhibition of these target genes via Srebf1 and Srebf 2 downregulation by intervening salidroside, a type of phenylethanoid glycoside found in Rhodiola and Ligustrum plant species, has been demonstrated by Song et al. (2021).Salidroside also inhibits the acetyl-CoA-producing enzyme encoded by the ATP citrate lyase gene (Acly) and leads to an increase in the production of 3-hydroxybutyrate (a ketone body) due to fatty acid degradation in apoE-deficient mice, resulting in enhanced glycerolipid and glycerophospholipid metabolism. 12Fuzhuan brick tea, a postfermented tea, has been produced in China since 1860 and involves fungal fermentation, which could lead to an increment in organic acids and tea pigments; its inhibitory effects on obesity in different models have been previously reported. 13Furthermore, Fuzhuan brick tea has recently been found to reduce fat

Journal of Agricultural and Food Chemistry
storage by suppressing the SREBP/MDT-15/FAT-2 pathway under normal cholesterol intake but may promote fat storage via the same pathway under a high-cholesterol dietary intake in the C. elegans model. 13Since SREBPs are important in cholesterol synthesis and metabolism, they are considered a major target for suppressing cholesterol biosynthesis.

3-Hydroxy-3-methylglutaryl-CoA Synthase 1 (HMGCS1) and 3-Hydroxy-3-methylglutaryl-coenzyme A Reductase (HMGCR).
HMGCS1 is a key enzyme in free cholesterol synthesis and is regulated by sterol regulatory element-binding protein 2 (SREBP2) and signal transducer and activator of transcription 1 (STAT1). 14The inhibition of HMGCS1 by direct binding with phytochemicals could thus be considered a new strategy in cholesterol reduction.Ursolic acid, a phytochemical widely found in medicinal herbs such as rosemary and thyme, has been previously shown to alleviate hypercholesterolemia and cardiovascular disease. 15 Ma et al.  (2022) revealed that ursolic acid could covalently bind to the thiol of Cys129 in HMGCS1, resulting in the inhibition of its catalytic activity and reduced precursor generation in cholesterol biosynthesis. 15Similarly, ligustilide, a natural phthalide derivative found in Ligusticum striatum and Angelica sinensis, also exhibits a regulatory effect on lipid metabolism by irreversibly binding to Cys129 of HMGCS1 after its metabolization to the intermediate 6,7-epoxyligustilide. 16 HMGCR is the main target of statins, a class of cholesterollowering drugs.HMGCS converts acetoacetyl-CoA into 3hydroxy-3-methylglutaryl-CoA (HMG-CoA), which is sub-sequently converted into mevalonate by HMGCR.Research has demonstrated that crocin, chlorogenic acid, geniposide, and quercetin intervention could reduce lipid deposition in a cell model by promoting the expression of cholesterolmetabolism-related genes and the suppression of SREBP2; furthermore, the combination of these four compounds exhibits a synergistic effect in inhibiting HMGCR expression. 17n addition, the drug simvastatin, used as a positive control, could not induce ATP-binding cassette (ABC) A1 and AMPactivated protein kinase (AMPK) α mRNA levels, as the natural compounds did.Unlike already-developed drugs, phytochemicals may exert their regulatory effects on multiple targets; therefore, further studies are needed to confirm the effect of phytochemicals as potential drugs for cholesterol dysregulation.
2.1.3.Farnesyl-Diphosphate Farnesyltransferase 1 (FDFT1, Squalene Synthase).In addition to HMGCS1, other enzymes involved in cholesterol synthesis and regulated by SREBPs could be inhibited by phytochemicals.Research has shown that bavachinin, a natural compound isolated from the traditional Chinese medicine Fructus Psoraleae, exhibits lipid-lowering effects by acting as a peroxisome proliferatoractivated receptor (PPAR) agonist and inhibits squalene synthase/FDFT1 via the suppression of the protein kinase B (Akt)/mammalian target of rapamycin (mTOR)/SREBP2 signaling pathway. 18.2.Phytochemical Inhibition of Cancer-Related Effects of Cholesterol Biosynthesis.In addition to Table 1.continued

Journal of Agricultural and Food Chemistry
inhibiting metabolic diseases, blocking cholesterol biosynthesis could serve as a potential cancer prevention strategy.Gypenosides, the major constituent of Gynostemma pentaphyllum, disrupt cholesterol biosynthesis via HMGCS1 inhibition, consequently suppressing the proliferation and migration of the Huh-7 and Hep3B cell lines. 19The suppressive effect of phytochemicals on cholesterol biosynthesis has been suggested in a colorectal cancer model.The callus extract of Hibiscus syriacus L., which is rich in fumaric acid and denatonium benzoate, was found to mediate Notch signaling, which subsequently suppressed cholesterol synthesis. 20Moreover, the proteins related to cholesterol biosynthesis, including HMGCR, FDFT1, squalene epoxidase (SQLE), and lanosterol synthase (LSS), were downregulated due to a reduction in Notch signaling and the activation of AMPK signaling.Notably, the callus extracts with the candidate compounds, such as fumaric acid, succinic acid, and denatonium benzoate, showed no toxicity toward normal cells; therefore, they could serve as potential candidates for cholesterol biosynthesis inhibition.
Wang et al. ( 2023) demonstrated that the extracellular matrix protein 1 (ECM1), elevated levels of which have been identified in malignant epithelial tumors, could regulate cholesterol biosynthesis and consequently trigger angiogenesis and cancer malignancy. 21Notably, the herb pair of Citri Reticulatae Pericarpium and Reynoutria japonica Houtt.have exhibited an inhibitory effect on target genes involved in the cholesterol metabolic process, including HMGCS1, FDFT1, mevalonate diphosphate decarboxylase (MVD), SQLE, and HMGCR, via ECM1 suppression.Studies have also shown that KRAS mutations in lung adenocarcinomas stop cholesterol efflux, thereby promoting tumor growth.Therefore, cholesterol removal via the promotion of the cholesterol efflux pathway could present a potential strategy to limit tumor growth. 22.3.Promotion of Cholesterol Biosynthesis by Phytochemicals.Some phytochemicals have been shown to promote cholesterol biosynthesis.For example, Chen et al. (2023) demonstrated that polyphyllin D, a steroidal saponin found in the rhizomes of Paris polyphylla, could induce cholesterol biosynthesis in liver cancer cells, with upregulation of the proteins involved, including the low-density lipoprotein receptor (LDLR, 22.17-fold change), RUN and FYVE domaincontaining protein 1 (RUFY1, 5.92-fold change), PPAL and HMGCR (an approximately 4-fold change), etc. 23 Although polyphyllin D has shown a disruptive effect on cholesterol biosynthesis in liver cancer cells, the regulatory effect on the cholesterol biosynthesis of normal hepatic cells should be addressed in future studies.

Effect of Phytochemicals on Cholesterol Uptake and Transport. As commonly known, low-density lipoprotein-cholesterol (LDL-C
) is compulsory for cholesterol transport to various organs, though the LDL-C level presents a causal risk factor for atherosclerosis. 24Regulating cholesterol uptake and excretion to maintain host cholesterol homeostasis could represent a strategy for the pharmacological treatment of related diseases.This section discusses the phytochemicals that could potentially regulate cholesterol uptake (Figure 2) and transport.
2.4.1.Cholesterol Uptake.LDLR is negatively regulated by PCSK9, and its reduction in the liver could lead to increased plasma LDL cholesterol, leading, in turn, to hypercholesterolemia. 25 Therefore, the regulation of these proteins by inducing LDLR expression with PCSK9 inhibition could potentially control hepatic cholesterol uptake.
The methanol extract (50% methanol) of the bergamot fruit contains several glycosidic and nonglycosidic flavonoids, with naringenin-7-O-rutinoside and apigenin-6,8-C-glucoside exhibiting a greater effect on inducing LDLR expression among the compared compounds. 26Moreover, bergamot peel extract can reduce PCSK9 and its transcription factor, hepatocyte nuclear factor 1 (HNF1-α), resulting in LDL uptake in Huh7 cell lines.Other flavonoids, such as a major compound isolated from the Gentiana veitchiorum flower, have exhibited an ameliorative effect on the liver by promoting the LDLR-lecithin-cholesterol acyltransferase (LCAT) signaling pathway and, thereby, preventing hepatic oxidative damage. 27Notably, PCSK9 is the transcriptional target of SREBP2 and subsequently reduces LDLR levels; AMPK activation could be responsible for the reduction of SREBP2/PCSK9, thus preserving LDLR expression. 28In another study, as a key receptor able to facilitate PCSK9 transport, sortilin was inhibited by crocetin and exerted a hypocholesterolemic effect via the preservation of LDLR expression in the HepG2 cell line. 29s previously mentioned, SREBP2 inhibition is a potential strategy to suppress cholesterol biosynthesis.However, some studies have revealed that its activation could be beneficial.For instance, drugs such as simvastatin can promote the levels of SREBP2 and LDLR to mitigate hyperlipidemia. 30Both PCSK9 and LDLR are transcriptionally regulated by SREBP2 to control lipid metabolism and maintain lipid homeostasis; therefore, the regulatory effect of phytochemicals on SREBP2, which may consequently affect PCSK9 and LDLR levels, should be evaluated.A study in 2021 demonstrated the regulatory effect of gypenosides on hepatic LDL, where no significant effect was observed in the expression of SREBP2, while the LDLR expression was upregulated, and the PCSK9 level was suppressed. 31The authors suggested that the regulation of PCSK9 and LDLR could be stimulated by SREBP-independent pathways.
Notably, scholars have indicated that berberine, a cholesterol-lowering drug, can selectively facilitate the growth of Blautia producta in mice guts, where the enrichment could lead to an LDL-lowering effect via LDLR promotion. 32The effect was further proven by treating the HepG2 cell line with Blautia producta, resulting in a LDL absorption.These findings indicate the multiple effects of phytochemicals on cholesterol uptake, though further evidence is required in this regard.

Cholesterol Transport.
Reverse cholesterol transport (RCT) involves the collection of excess cholesterol by HDL and its delivery to the liver, where it is broken down and excreted. 33ABC transporter proteins from the A subfamily are responsible for mediating RCT; on the other hand, those from the G subfamily are responsible for cholesterol efflux, with the most well-known member being the ABCG5-ABCG8 heterodimer. 34ome phytochemicals from Chinese traditional herbs employ this mechanism.For instance, the Qishen Yiqi Chinese herb pill, consisting of astragaloside IV, salvianolic acid B, and notoginsenoside R1, increases ABCA1 expression via the PPAR-γ/liver X receptor (LXRα/β) pathway to promote the RCT signaling pathway, leading to the attenuation of atherosclerotic lesions. 35In 2023, researchers reported that the Huazhuotongmai decoction (traditional Chinese medicine), containing up to 30 different bioactive compounds, promoted RCT by upregulating the expression of ABCA1, scavenger receptor class B type I (SR-BI), and PPAR-γ in rabbits fed a high-fat diet, resulting in an antiatherosclerotic effect. 36In addition, the Jieduquyuziyin prescription (Chinese medicine) promotes cholesterol efflux via a similar pathway in mice livers. 37Based on the above-mentioned studies, RCT has recently been identified as a potential mechanism of defense against atherosclerosis progression.
Gypenoside XVII, a natural compound isolated from the traditional herbal medicine Gynostemma pentaphyllum Makino, has been shown to promote the expression of ABCA1 and ABCG1 via the inhibition of histone deacetylase 9 (HDAC9) expression by miR-182-5p in lipid-loaded macrophages. 38In addition, activation of miR-182-5p/HDAC9 by gypenoside XVII to promote cholesterol efflux potentially prevented oxidized low-density lipoprotein (ox-LDL)-induced macrophages and avoided atherosclerotic plaque formation.Similarly, fargesin, a neolignan isolated from Magnolia plants, exhibited a promoting effect on RCT in both apoE −/− mice and ox-LDLinduced macrophages. 39Notably, fargesin upregulated ABCA1 and ABCG1 via the CCAAT/enhancer binding protein (CEBP)α/LXRα pathway and specifically increased the level of phosphorylation of CEBPα in Ser21 in THP1-derived macrophages.
Polydatin, also known as 3,4′,5-trihydroxystilbene-3-βmono-D-glucoside, is a bioactive compound detected in various plants, such as grapes and peanuts.By inducing cholesterol efflux via ABCA1, ABCG1, and SR-BI in aortic macrophages, polydatin reduced the formation of foam cells in the aorta. 40In addition, it promoted the expression of ABCG5/G8 and CYP7A1 for the secretion of cholesterol.Yuan et al. (2020)  showed that phytosterols (ergosterol, stigmasterol, β-sitosterol, campesterol, ergosterol acetate, and stellasterol) regulate the uptake and efflux of cholesterol.Specifically, NPC1-like intracellular cholesterol transporter 1 (NPC1L1) was inhibited to reduce the absorption of cholesterol, while ABCG5/G8 levels were enhanced to increase its excretion. 41Although the cholesterol-lowering effect via multiple mechanisms of phytosterols has been previously reviewed, 42 the authors of this study suggested that the types and positions of functional groups on the phytosterols might contribute to their cholesterol-lowering effects.On the other hand, a recent study has revealed that phytosterol ferulate, a compound naturally present in certain grains (albeit in low amounts), demonstrated a lipid-lowering effect that proved to be more effective than merely combining phytosterols and ferulic acid. 43ower levels of serum and hepatic TC were observed in obese mice that were supplemented with the sample; however, the underlying mechanism still needs further clarification.Moreover, several structural modified plant sterols have been identified which have high potential to be developed into cholesterol controlling supplements. 44ertain other members of the ABC transporter family play an important role in cholesterol transport in various tissues; the regulation of these transporters and their roles in disease amelioration should be investigated in future studies.

ASSOCIATION OF BILE ACIDS WITH CHOLESTEROL HOMEOSTASIS
Synthesized in the liver from cholesterol, bile acids are conserved molecules and are essential for lipid homeostasis.Bile acid synthesis is a multistep process involving up to 17 enzymes located in different cellular organelles.The liver is responsible for the catabolism of cholesterol into bile acids since it contains all the required enzymes for the process. 45Bile acid metabolism is crucial in the regulation of lipid metabolism and the maintenance of cholesterol homeostasis (Figure 3).

Effect of Phytochemicals on Bile Acid Homeostasis.
Studies have demonstrated that the regulatory effect of phytochemicals on enzymes involved in the hepatic synthesis and colonic metabolism of bile acids can be used to prevent the conditions caused by excess cholesterol.The application of phytochemicals to promote bile acid synthesis, metabolism, and excretion and reduce hepatic cholesterol levels has thus been suggested as a potential strategy.Additionally, preventing the reabsorption of bile acids from the ileum should be considered.
Saikosaponins, oleanane derivatives found in Radix Bupleuri, exhibited a modulatory effect on cholesterol clearance in a recent study; cholesterol efflux was promoted by significant upregulation of the genes related to cholesterol transportation, such as Abca6, Apof, Npc1, Pdzk1, Ttc39b, and Scarb1.Furthermore, the rate-limiting enzymes responsible for bile acid biosynthesis and cholesterol sulfation, including cyp7a1, hsd3b7, and sult2b1, were upregulated by saikosaponin intervention. 46These findings indicate the ameliorative effect of saikosaponins on hepatic steatosis, partly via promotion of cholesterol clearance through cholesterol efflux and bile acid synthesis.Geniposide, mentioned earlier as promoting cholesterol metabolism, was also shown to induce bile acid synthesis and excretion.Both the protein and mRNA levels of CYP7a1, CYP27a1, CYP7b1, and CYP8b1 were upregulated in response to geniposide treatment, leading to hepatic bile acid synthesis via the consumption of hepatic cholesterol. 47he farnesoid X receptor (FXR) is the most important nuclear receptor for maintaining bile acid homeostasis, playing a specific role in suppressing bile acid synthesis and promoting its enterohepatic circulation. 48FXR can inhibit the synthesis of bile acid via its negative feedback regulation, but geniposide has been found to inhibit the hepatic FXR level, accompanied by its downstream targets, bZip Maf transcription factor (MAFG) and tyrosine phosphatase (SHP).As further evidence, increased levels of bile acids in urine and feces suggest that bile acid excretion is promoted by geniposide intervention.In addition, the reduction in ileum FXR, ileal bile acid-binding protein (I-BABP), and apical sodium-dependent bile acid transporter (ASBT) suggests that geniposide intervention reduces bile acid reabsorption as a form of protection against these acids. 47he studies mentioned in this section suggest that the maintenance of bile acid levels is an effective strategy to prevent cholesterol-related diseases.Promoted bile acid synthesis should be accompanied by its increased excretion and/or the reduced resorption of colonic bile acid to prevent bile acid accumulation in the liver and simultaneously clear the level of hepatic cholesterol.

Effect of Phytochemicals on Bile Acid
Recomposition and Recovery/Reabsorption.Bile acid is known to facilitate the digestion and absorption of lipids, and more importantly, regulate cholesterol homeostasis. 49Therefore, blindly promoting bile acid excretion could have a negative effect on the host.The strategic recomposition of the bile acid pool and promotion of bile acid reabsorption can, thus, maintain the host's health.
Although the above-mentioned study reveals geniposide's inhibitory effect on hepatic FXR in mice fed a high-fat diet, its promoting effect on FXR has been observed in bile duct ligation-induced cholestasis in mice models.Research has shown that the binding of geniposide to sirtuin 1 (SIRT1) increases the deacetylation of hepatic FXR and restores bile acid profiles/contents, especially those of chenodeoxycholic acid (CDCA), tauroursodeoxycholic acid (TUDCA), and taurochenodeoxycholic acid (TCDCA), ultimately ameliorating liver fibrosis. 50Therefore, phytochemicals might exhibit distinct effects under different disease conditions, and their benefits should be further clarified.Li et al. (2023) have reported similar results, demonstrating that vine tea extract restores hepatic and ileum FXR and ASBT�inhibiting bile acid synthesis and promoting its transport and reabsorption� and can serve as a means to improve hepatic injury. 51Scholars have also reported Forsythiaside A, the main active compound of Forsythia suspensa (Thunb.)Vahl, has a similar effect on hepatic bile acid synthesis in a CCl 4 -induced liver fibrosis mouse model.Forsythiaside A significantly reversed the suppression of the mRNA related to hepatic FXR-mediated bile acid regulation�CYP7A1, SHP, and liver receptor homologue-1 (LRH-1)�and transport−bile salt export pump (BSEP), multidrug resistance protein 4 (MRP4), Na +taurocholate cotransporting polypeptide (NTCP), and organic anion transporting polypeptide-1 (OATP-1)�to restore the normal levels. 52Forsythiaside A intervention also reversed all CCl 4 -induced abnormal changes in bile acid levels.
The major characteristic of cholestasis is the accumulation of hepatic bile, and the activation of FXR to impede bile acid synthesis and promote its transportation is considered to be a promising strategy for maintaining bile acid homeostasis.Researchers have reported that Dolomiaea souliei (Franch.)C. Shih, a herbal medicine with costunolide as a major constituent, structurally interacts with FXR to prevent cholestasis. 53The binding of the compound to FXR significantly activated the FXR/SHP pathway; inhibited CYP7A1 and CYP27A1 to control the bile acid synthetic process; and upregulated BSEP, MRP2, and NTCP to promote bile acid transport, consequently ameliorating bile accumulation in the liver.
G protein-coupled bile acid receptor 1 (TGR5) is a transmembrane G-protein-coupled receptor (GPCR) for bile acids.Other than FXR, TGR5 has also been identified as a regulatory target of bile acid and may facilitate the occurrence of lipolysis and energy consumption. 54Research has indicated that red ginseng extract promotes ASBT membrane localization via TGR5 activation, resulting in bile acid uptake in mice intestines. 55The effect of red ginseng supplementation was reflected in the serum, white adipose tissue, and ileum bile acid profiles, although no significant changes in the composition of hepatic bile acid were observed.In another study, ginsenoside compound K activated TGR5 in the L-cells of the intestinal epithelium, possibly via the gut microbiota− bile acid axis. 56The intervention of compound K significantly facilitated the growth of Akkermansia, Lactobacillaceae, Lachnospiraceae, and Ruminococcaceae and simultaneously reduced the abundance of Bacteroidaceae and Enterococcaceae.Moreover, the deoxycholic acid (DCA) and lithocholic acid (LCA) levels increased significantly after treatment with compound K, which subsequently activated TGR5.In addition, the upregulation of CYP7B1 and CYP27A1 and downregulation of CYP8B1 occurred due to changes in the bile acid composition.
Both the activation of FXR and TGR5 and the inhibition of FXR by phytochemicals promote the maintenance of bile acid homeostasis.These results indicate that removing their expression might lead to negative effects.Therefore, phytochemicals and their regulatory effects are preferred over drugs in preventing side effects.

RELATION OF CHOLESTEROL REGULATION TO BILE ACIDS VIA GUT MICROBIAL MODULATION
Many reports have indicated the correlation between the microbiome and the incidence of cardiovascular disease. 57,58esearch has shown that the depletion of gut microbiota increases intestinal cholesterol uptake and cholesterol biosynthesis in the liver. 59In another study, the microbial conversion of cholesterol into coprostanol beneficially reduced intestinal and serum cholesterol levels, revealing the crucial role of gut microbiota. 60It is also understood that around 95% of bile acids are reabsorbed and recycled via enterohepatic circulation, while the remaining amount enters the colon and is dehydroxylated into DCA and LCA by gut microbes. 61FXR is activated by ligands, especially hydrophobic bile acids such as CDCA, followed by LCA and DCA (to a similar extent) and cholic acid (CA; with a lower potency). 62The activation of FXR is negatively correlated to CYP7A1, CYP8B1, CYP27A1, and CYP7B1, which are involved in inducing bile acid synthesis. 62Therefore, the colonic bile acid composition significantly influences the rate of hepatic bile acid synthesis.

Effect of Phytochemicals on
Cholesterol/Bile Acid-Related Genes through Gut Microbiota.Moringa-Fu brick tea is produced via cofermentation technology combining the dark tea and moringa leaves, and its health benefits in regard to metabolic disorders, arising from its polyphenol and theabrownin content, have been previously reported. 63Recent studies have also indicated its effectiveness in reducing hepatic cholesterol via the regulation of bile acid metabolism (including CYP7A1 and CYP27A1 via the reduction of intestinal FXR); notably, the abundance of gut microbes associated with bile salt hydrolase (BSH) activity was regulated by Moringa-Fu brick tea extract. 63Some gut microbes, including Akkermansia, Clostridium, Bacteroides, Roseburia, Parabacteroides, and Prevotella, are positively correlated with serum high-density lipoprotein-cholesterol (HDL-C) and hepatic expressions of CYP7A1 and CYP27A1.The effect of chin brick tea, Camellia sinensis (L.) Kuntze, on modulating intestinal flora and, consequently, altering the composition of bile acid has also been recently reported.Polyphenols in the aqueous extract of chin brick tea (rich in catechin-like compounds) significantly increased the primary/secondary bile acid ratio.Specifically, Bacteroides positively correlated to nonalcoholic fatty liver disease (NAFLD) were positively correlated with DCA and ωmuricholic acid (ωMCA), while Lactobacillus exhibited a negative correlation with these, serum LDL-C/HDL-C, and the hepatic total cholesterol (TC). 64These studies suggest the modulatory effect of teas on gut microbial composition, which can consequently affect bile acid synthesis and metabolism.Therefore, further investigations are required to clarify the correlation between the composition of the bile acid and the abundance of specific gut microbes.
In addition to tea, other plant-based supplements display regulatory effects on the gut microbiota.Flaxseed powder, a commercial source of dietary α-linolenic acid, exhibits an ameliorative effect on high-fat-diet (HFD)-induced NAFLD, possibly by regulating gut microbiota and bile acid metabolism.Increases in the relative abundances of Parasutterella, Lachnospiraceae_NK4A136_group, and [Eubacterium]_xylano-philum_group have been reported in flaxseed-supplemented groups, accompanied by repression in the relative abundances of Coriobacteriaceae_UCG-002, Erysipelatoclostridium, and [Eubacterium]_f issicatena_group.A positive correlation has been reported between some bile acid metabolites and Erysipelatoclostridium and [Eubacterium]_f issicatena_group. 65otably, changes in intestinal bile acid have led to the activation of FXR and resulted in a decrease in hepatic cyp7a1 and cyp8b1, which control bile acid synthesis.
Disruptions to bile acid synthesis and metabolism may contribute to liver disease due to HFD intake.In an ovariectomized-induced dyslipidemia rat model, Radix Angelica dahuricae extract was found to interact with the proteins related to bile acid signaling pathways, including FXR, LXR, and PPARα.Radix Angelica dahuricae is a Chinese traditional medicine with furanocoumarins as active compounds, including imperatorin, isoimperatorin, oxypeucedanin hydrate, and oxypeucedanin; 66 its extract increased BSH levels in the liver and cecum and resulted in the recomposition of bile acid.Significantly elevated levels of Lactobacillus reuteri, Ruminococcus bromii, and Parabacteroides distasonis were also identified in the group treated with Radix Angelica dahuricae extract. 66enerally, the bile acid pool is composed of both primary bile acids, including CA and CDCA, and secondary bile acids, such as DCA and LCA.The abundance of Lactobacillus reuteri was positively correlated to CA levels but negatively correlated to taurohyodeoxycholic acid (THDCA).In addition, Ruminococ- Another study demonstrated the modulatory effect of a traditional Chinese formula comprising several traditional herbs on gut microbiota and revealed the correlation between some gut microbes and bile acids.Akkermansia, Allobaculum, Bilophila, Clostridium, and Lactobacillus were positively correlated to hepatic unconjugated bile acids and negatively correlated to conjugated ones; these gut microbes were positively correlated to the primary bile acids in feces and negatively correlated to secondary ones.Notably, Coprococcus and Halomonas showed opposite correlations with Akkermansia, Allobaculum, Bilophila, Clostridium, and Lactobacillus. 67.2.Effect of Phytochemicals on Bile Acid Profiles through Gut Microbes.Berberine, an alkaloid isolated from the rhizome of Coptis chinensis, has recently been reported to improve gut microbiota depletion in a mouse colitis model by facilitating the growth of the Lactobacillus and Roseburia genera; this resulted in an increase in the colonic levels of CDCA and DCA, which activated colonic FXR and inhibited hepatic CYP7A1, leading to the control of bile acid synthesis.Reports have indicated that FXR/fibroblast growth factor 15 (FGF15) activation occurs via the intervention of Tripterygium hypoglaucum (Levl.)Hutch extract, a source of diterpenoids; the activation leads to a reduction in free bile acids and an increase in conjugated bile acids. 68The changes in the bile acid composition are listed in Table 2.

Recomposition of Gut Microbial Profiles
Promoting Bile Acid Synthesis.The above-mentioned studies demonstrate the suppression of bile acid synthesis to prevent metabolic diseases and avoid damage by bile acids.In contrast, other studies have suggested the enhancement of bile acid synthesis as a strategy to reduce hepatic cholesterol levels.The phytosterol stigmasterol, a natural steroid, has been widely studied and exhibits antihyperlipidemic and cholesterollowering properties.Stigmasterol intervention has not only reduced hepatic free cholesterol and 25-hydroxycholesterol levels but has also promoted alternative bile acid synthetic pathways (cyp27a1, cyp7b1, and bsep levels) and, subsequently, enhanced the fecal excretion of bile acids. 69Specifically, the colonic levels of hydrophilic bile acids (ursodeoxycholic acid [UDCA], CA, α-MCA, β-MCA, and CDCA) were significantly increased in the stigmasterol intervention group, while hydrophobic bile acids, such as DCA and LCA, were notably reduced, facilitating the excretion of bile acids via feces.The alterations in bile acids could be due to the reshaping of the gut microbiota by stigmasterol, which can significantly increase the Chao1 index and modulate the gut microbial compositions.The correlations between the different bile acids and gut microbes reviewed in this study are presented in Table 2.

SUMMARY
Cholesterol is pivotal in the biological functioning of the body, but the Westernization of dietary patterns has promoted the incidence of metabolic diseases related to cholesterol dysregulation.Although a plethora of studies have focused on the potential effects of dietary compounds on cholesterolrelated diseases, the outcomes are subtle.Studies on the effect of traditional Chinese medicines have led to some striking results, although there is a lack of sufficient scientific evidence to confirm and substantiate their significance in terms of disease improvement.Therefore, further studies should be conducted to identify the primary components responsible for Table 2. continued

Journal of Agricultural and Food Chemistry
the effects attributed to traditional Chinese medicines.Additionally, it is crucial to investigate whether these effects result from a synergistic interaction among the multiple beneficial components.More importantly, the specific underlying mechanisms that could be either promoted or inhibited by these components need to be further elucidated.Nevertheless, these phytochemicals could still serve as promising candidates to overcome the shortcomings of already-developed drugs (i.e., their side effects).Cholesterol remains important in the functioning of the body and thus should not be completely eliminated from the body by drugs; therefore, retuning and improving cholesterol levels could represent better strategies, though these present challenges, to some extent, for developed drugs.Furthermore, complications arising from gut microbiota, which, to a large extent, are modulated but not manipulated, because gut microbiota are vulnerable to change.Thus, perfectly improving cholesterol-related diseases with drugs is virtually impossible.In summary, studies on dietary compounds for cholesterol regulation cannot be neglected, and the development of therapeutic adjuvants for cholesterol dysregulation can be addressed in future studies.

Table 1 .
Effect of Phytochemicals on Cholesterol Metabolism in Different Disease Models

Table 2 .
Effect of Phytochemicals on Bile Acid and Gut Microbial Composition

Journal of and Food Chemistry pubs.acs.org/JAFC Review
cus bromii showed a negative relationship with taurocholic acid (TCA), TUDCA, and THDCA but promoted the level of primary bile acids (CDCA) in humans.